The overall goal of this study is to elucidate the role of the apoptosis-related protease, caspase-2, in male age-dependent osteoporosis. Our lab has demonstrated that mice deficient in caspase-2 not only have decreased maximum life span, but also show features of premature aging compared to age-matched wild-type mice. Examples of aging-related traits seen in caspase-2 deficient mice include impaired hair regrowth ability, decreased body fat content, increased levels of protein oxidation in the liver, and importantly, severe bone loss. Specifically, we found decreased trabecular bone volume, an increased level of the clinical osteoporosis marker, deoxypyridinoline (DPD), in urine, and increased numbers of osteoclasts, cells responsible for bone resorption. These results suggest that caspase-2 deficiency may prevent osteoclast spontaneous apoptosis, which is induced by oxidative stress. Additionally, we did not observe any compensatory increase in bone formation or osteoblast number, an issue also observed in human osteoporosis. Because of these findings, we hypothesize that caspase-2 plays a critical role in osteoclast apoptosis induced by oxidative stress. Furthermore, because the aging process has been linked to increased oxidative stress levels in cells, we hypothesize that capase-2 is critical in mediating apoptosis of aging osteoclasts, leading to inhibition of excess bone resorption. Our first specific aim will determine if caspase-2 normally induces oxidative stress-dependent apoptosis in osteoclasts as the organism ages. We will determine the levels of oxidative stress markers such as reactive oxidative species (ROS), glutathione (GSH), and NADPH in young and old caspase-2 knockout mice and compare them to levels seen in osteoclasts from age-matched wild-type mice. Additionally, we will investigate the role of oxidative stress in inducing spontaneous apoptosis of aging osteoclasts isolated from caspase-2 knockout and wild-type mice. Our second aim involves determining the role of caspase-2 and aging in osteoblast-osteoclast interactions. This will be accomplished by measuring the expression level of different factors that have been shown to affect osteoblastogenesis (Wnt10b, BMP6, and PDGF BB) or osteoclastogenesis (RANKL, OPG, and M-CSF). These factors will be measured from bone marrow cultures, containing osteoblasts and osteoclasts, generated from bone in young and old caspase-2 knockout and wild- type mice. Furthermore, the factor-rich media from the co-culture will be used as a conditioned media to study osteoblastogenesis and osteoclastogenesis in precursor cells. Understanding the function of caspase-2 in age-dependent osteoporosis, on one hand, and showing how changes in osteoblast-osteoclast interactions can lead to severe bone resorption, on the other, will lead to putative identification of molecular targets that may develop into new and valuable anti-osteoporosis therapies.